Both immunotoxins and TNF have been demonstrated to produce antitumor activity in mammals. As with any treatment using potent chemotherapeutic agents, toxic side effects may occur. These effects may be minimized if the dosage levels are kept as low as possible. By utilizing a combination of TNF and a suitable immunotoxin the dose level of each component can be reduced, thus preventing the toxicity which might otherwise be encountered at the ordinarily used dosaqe levels.
TNF itself is, of course, noted for its ability to hamper tumor growth and kill tumor cells. An extensive literature on TNF per se exists, and the gene encoding native TNF proteins has been cloned and expressed.
Tumor necrosis factor (TNF) was first described by Carswell et al, Proc Natl Acad Sci USA (1975), 72:3666-3670. as an endotoxin-induced serum factor which causes necrosis of chemically transformed tumor cells when growing in mice. Purified preparations of murine TNF have been tested against murine and human cell lines in vitro: Haranaka, K., and Satomi, N., Japan J Exp Med (1981) 51:191. In contrast to normal cells, tumor cell lines from both species were susceptible to the cytotoxic activity of the mouse TNF. Furthermore, the murine TNF was reported to be toxic against both humanand mouse-transplanted tumors in nude mice. See Haranaka. K.. et al, Int J Cancer (1984) 34:263-267. Human TNF is also known to be cytotoxic to neoplastic cells, and has been produced in recombinant form. See Pennica et al. Nature (1984) 312:724-729; Shirai et al, Nature (1985) 313:803-806; Wanq et al, Science (1985) 228:149-154.
The cloning of rabbit TNF is disclosed in EP No. 146,026. published 26 June 1985 (Dainippon Pharmaceutical Co., Ltd.) and EP No. 148,311, published 17 July 1985 (Asahi Kasei Kogyo Kabushiki). The cloning of human TNF having 151 and 155 amino acids (2 and 6 less than the human native form) is disclosed in EP No. 155,549, published 25 Sept. 1985 (Dainippon Pharmaceutical Co., Ltd.), and human TNF having 155 amino acids is disclosed in EP No. 158,286, published 16 Oct. 1985 (Asahi Kasei Koqyo Kabushiki Kaisha) corresponding to GB No. 2,158,829A, published 20 Nov. 1985. The cloning of mature TNF (157 amino acids) and various modified forms (muteins) thereof is disclosed in EP No. 168,214, published 15 Jan. 1986 (Genentech) and PCT U.S. Pat. No. 85/01921, filed 3 Oct. 1985. published Apr. 1986 (Cetus Corporation). The latter, PCT No. 85/01921, corresponds to U.S. Ser. No. 760,661 filed 30 July 1985, now U.S. Pat. No. 4,677,063 the disclosure of which is incorporated herein by reference.
The literature on potential immunotoxins against tumors is also extensive. In most constructions, the tumor specificity is provided by an antibody or fragment thereof which is immunoreactive with tumor surface antigens. Both polyclonal sera raised against tumor antigens and monoclonal antibody preparations have been employed. The remainder of the immunotoxin is a cytotoxic moiety which can be chosen from a broad range of candidates.
Antisera and monoclonal preparations against a variety of tumors have been prepared and are known in the art. In particular, murine monoclonal antibodies that bind selectively to human breast cancer cells have been prepared. When conjugated to ricin A chain to form an immunotoxin, these antibodies exhibit a tissue culture inhibitory dose which results in 50% of control (untreated) protein synthesis (TCID 50%) of less than about 10 nM against at least one of MCF-7, CAMA- 1, SKBR-3. or BT-20 cells. These antibodies are described more fully in EPC Patent Publication No. 153,114, published 28 Aug. 1985,the disclosure of which is incorporated herein by reference.
Similar murine monoclonal antibodies have been used for imaging. These antibodies do not bind to blood cells, and have a breast tumor binding range of at least 0.25 (i.e., they bind to at least 25% of breast tumors tested). These antibodies include most of those described above and are described in copendinq U.S. application Ser. No. 786,948, filed 11 Oct. 1985, assigned to the same assignee and incorporated herein by reference.
Combination chemotherapy in general using two or more anti-cancer drugs to treat malignant tumors in humans is currently in use in research and in the clinic. The anti-cancer druqs may be antimetabolites, alkylating agents, antibiotics, general poisons. etc. Combinations of drugs are administered in an attempt to obtain a synergistic cytotoxic effect on most cancers, e.g., carcinomas. melanomas, lymphomas and sarcomas and to reduce or eliminate emergence of drug-resistant cells and reduce side effects of each drug. Dr. Talmadge of the Preclinical Screening Lab, BRMP reported in 1986 the augmented effect of using TNF and .gamma.-IFN to treat metastatic disease in mice. EP publication No. 131,789 published 23 Jan. 85 (Sloan-Kettering Institute for Cancer Research) and EP No. 168,214 published 15 Jan. 85 (Genentech) disclose the synergistic effect of the TNF and .lambda.-IFN to treat various tumors in mice. However, applicants are unaware of any demonstration of synergistic effects against tumors by combinations of suitable immunotoxins with TNF.